Alcohol addiction is a chronically relapsing disorder characterized by compulsive drug-seeking and use. A major factor contributing to this profile of behavior is the process of associative learning whereby environmental stimuli paired with alcohol consumption acquire incentive-motivational value. Little is known about the processes leading to the development of the compulsive-like nature of ethanol-seeking. The objective of this proposal is to elucidate these processes with emphasis on identifying neural substrates responsible for the distinctly compulsive nature of alcohol-seeking as opposed to those mediating behavior motivated by natural reward essential for survival, well being and healthy hedonic pursuits. To accomplish this, a novel behavioral model will be employed based on the observation that alcohol-seeking elicited by ethanol (EtOH) -associated contextual cues (conditioned reinstatement) shows remarkable persistence whereas the motivating effects of stimuli conditioned to highly potent natural reward (PNR) decay rapidly. This differential persistence of conditioned reinstatement will serve as the central model to dissociate compulsive- like from normal seeking behavior. Based on preliminary data, these studies will target several primary neural systems: The hypothalamic orexin/hypocretin (Orx/Hcrt) and CART (cocaine and amphetamine-related transcript) systems, as well as Group II metabotropic glutamate receptors. A second objective will be to identify novel neural systems that regulate compulsive ethanol-seeking. The research plan employs three converging approaches for detecting and verifying a role of specific brain sites and neural systems in the differential persistence of reward-seeking that characterizes reinstatement induced by EtOH-associated contextual cues vs. stimuli conditioned to PNR: Initially, (in SPECIFIC AIM I), a systematic neural mapping approach will be employed to differentiate brain sites activated by an EtOH cue from those responsive to a PNR cue, using inducible transcription factors (ITF) as markers. This strategy will be complemented in SPECIFIC AIM II by directly targeting the signaling systems about which specific hypotheses have been formulated based on preliminary data. Specifically, levels of Orx/Hcrt and CART immunoreactivity as well as mGlu2 and mGlu3 receptor expression will be determined to test whether a differential role of these signaling systems in seeking behavior induced by the EtOH vs. PNR cue can be traced to differential alterations within distinct brain sites. Additionally, both in the case of Orx/Hcrt and CART, and novel systems be identified, dual-labeling with one of the ITF markers showing activation will provide direct information on the phenotype of neurons showing activation in response to the EtOH vs. PNR cues. A role of signaling systems within specific brain sites implicated in the differential persistence of EtOH vs. PNR cue-induced reinstatement by results of Specific Aims I and II, then will be verified by site-specific pharmacological manipulations in SPECIFIC AIM III. The research plan has been developed with the objective of advancing understanding the biological basis of alcohol dependence, as well as of the neuroscience of motivated behavior, in general. PUBLIC HEALTH RELEVANCE: Alcohol addiction is a chronically relapsing disorder characterized by compulsive drug-seeking and use. The objective of this proposal is to identify neuroanatomical and neuropharmacological substrates responsible for the distinctly compulsive nature of alcohol-seeking as opposed to those mediating behavior motivated by natural reward essential for survival, well being and healthy hedonic pursuits. This research is expected to (a) advance understanding the biological basis of alcohol dependence, (b) reveal novel treatment targets for alcohol abuse and alcoholism, and (c) advance basic science understanding of normal and pathological goal- directed behavior, in general